Role of Stress-Response Protein gp96 in the Persistence of Rheumatoid Arthritis

描述（由申请人提供）：先天性和适应性免疫系统在类风湿性关节炎（RA）的发病机制中很重要。Toll样受体（TLR）对于包括T和B细胞活化的适应性免疫的产生是重要的。然而，TLR在RA的持续存在和破坏中的作用尚不清楚。我们最近证明，表达突变Fas受体（lpr）的小鼠，发展不太频繁和不太严重的胶原诱导性关节炎（CIA）。我们确定了一种新的机制，其中Fas-FasL相互作用的巨噬细胞（M&S），而不诱导凋亡，增强激活通过TLR 4途径。我们证明完整的Fas- FasL信号通路促进CIA的诱导并增强由TLR 4连接诱导的NF-：B活化和细胞因子表达。M&S和滑膜成纤维细胞在RA的发病机制中至关重要。RA关节中的M&s表达Fas和FasL，并与其他M&s和滑膜成纤维细胞密切接触。这些观察结果表明，在RA关节中，Fas-FasL相互作用，通过降低激活阈值，可以使滑膜M&S对TLR配体的激活敏感。我们的初步研究还表明，从RA关节分离的M&S与对照M&S相比，对微生物TLR 2和TLR 4配体的反应性增加，表明TLR信号传导在RA中可能很重要。我们的初步数据还表明，96-kDa的糖蛋白96（gp 96），内质网应激反应蛋白，是高度表达在RA关节，并激活M&S通过TLR 2和非凋亡的Fas-FasL相互作用促进M&激活gp 96。我们还证明，另一个潜在的TLR配体，热休克蛋白22，通过TLR 4激活，是丰富的RA滑液。总之，这些观察结果表明，M&s之间以及可能M&s和滑膜成纤维细胞之间的Fas-FasL相互作用可以使关节内的细胞局部对内源性TLR配体（例如gp 96或HSP 22）的活化敏感，从而导致自我持续的慢性炎症。本申请的首要假设是内源性TLR配体的局部释放促进关节炎从自限性疾病进展为慢性疾病，并且该进展通过Fas-FasL相互作用增强。具体而言，我们假设：1）使用gp 96和HSP 22，通过TLR结扎的RA滑膜M&s的活化将大于用正常M&s或来自其他形式的炎性关节炎（OIA）如银屑病关节炎患者的滑液的那些观察到的活化，而RA滑膜成纤维细胞的活化将大于用来自骨关节炎（OA）患者的滑膜成纤维细胞观察到的活化; 2）Fas-FasL相互作用是内源性TLR配体最大化TLR 2或TLR 4介导的活化所必需的; 3）在存在但不缺乏完整Fas信号传导的情况下，gp 96（或HSP 22）的表达将依赖于TLR的自限性关节炎转化为慢性进行性疾病，以及4）中和gp 96（和/或HSP 22）将改善CIA。 公共卫生相关性。风湿性关节炎影响至少1%的人口，并且有许多患者治疗不足。本研究中提出的工作旨在确定有助于类风湿关节炎发病机制的新机制，这将定义新的治疗靶点。

DESCRIPTION (provided by applicant): The innate and adaptive immune systems are important in the pathogenesis of rheumatoid arthritis (RA). Toll- like receptors (TLRs) are important for the generation of adaptive immunity, including T and B cell activation. However the role of TLRs in the persistence and destruction of RA is not known. We recently demonstrated that mice expressing a mutant Fas receptor (lpr), developed less frequent and less severe collagen induced arthritis (CIA). We identified a novel mechanism whereby Fas-FasL interactions on macrophages (M&s), while not inducing apoptosis, enhanced activation through the TLR4 pathway. We demonstrated that intact Fas- FasL signaling promoted the induction of CIA and enhanced the NF-:B activation and cytokine expression induced by TLR4 ligation. M&s and synovial fibroblasts are critical in the pathogenesis of RA. M&s in the RA joint, express both Fas and FasL and are in intimate contact with other M&s and with synovial fibroblasts. These observations suggest that, in the RA joint, Fas-FasL interactions, by lowering the threshold for activation, may sensitize synovial M&s to activation by TLR ligands. Our preliminary studies also demonstrate that M&s isolated from the RA joint demonstrate increased responsiveness to microbial TLR2 and TLR4 ligands compared to control M&s, suggesting that TLR signaling might be important in RA. Our preliminary data also demonstrate that the 96-kDa glycoprotein 96 (gp96), an endoplasmic reticulum stress response protein, is highly expressed in the RA joint, and activates M&s through TLR2 and that non-apoptotic Fas-FasL interactions promote M& activation by gp96. We also demonstrate that another potential TLR ligand, HSP22, which activates through TLR4, is abundant in RA synovial fluids. Together these observations suggest that Fas-FasL interactions between M&s, and perhaps between M&s and synovial fibroblasts, may sensitize cells locally within the joint to activation by endogenous TLR ligands, such gp96 or HSP22, resulting in a self- perpetuating chronic inflammation. The overriding hypothesis of this application is that the local release of endogenous TLR ligands promotes the progression of arthritis from a self-limited to a chronic disease, and that this progression is enhanced by Fas-FasL interactions. Specifically, we hypothesize that: 1) Employing gp96 and HSP22, the activation of RA synovial M&s through TLR ligation will be greater than observed with normal M&s or those from the synovial fluid of patients with other forms of inflammatory arthritis (OIA) such as psoriatic arthritis, while the activation of RA synovial fibroblasts will be greater than observed with synovial fibroblasts from patients with osteoarthritis (OA); 2) that Fas-FasL interactions are required for maximal TLR2 or TLR4-mediated activation by endogenous TLR ligands; 3) that the expression of gp96 (or HSP22) converts self-limited arthritis to chronic progressive disease, dependent upon TLRs, in the presence, but not the absence, of intact Fas signaling and that 4) that neutralization of gp96 (and/or HSP22) will ameliorate CIA. PUBLIC HEALTH RELEVANCE. Rheumatoid Arthritis affects at least 1% of the population and there are many patients for whom therapy is inadequate. The work proposed in this study is aimed at identifying novel mechanisms contributing to the pathogenesis of rheumatoid arthritis, which would define new therapeutic targets.